Full-mouth Treatment Approaches in Severe Chronic Periodontitis.

Study population and clinical examination

60 consecutive severe chronic periodontitis patients were recruited for the study.

The whole mouth clinical periodontal examination included measurement of probing depth, clinical attachment level and plaque index at 6 sites per tooth, except the third molars The papillary bleeding index was recorded for the entire oral cavity. Clinical periodontal parameters were recorded at baseline and 1 month, 3 months and 6 months after treatments. Alveolar bone loss were assessed using an digital panoromic radiograph in each participant.

The diagnosis of severe chronic periodontitis was based on clinical and radiographic diagnostic criteria proposed by 1999 International Workshop for a Classification of Periodontal Diseases and Conditions. These individuals had a minimum of four non-adjacent teeth with sites with PD ≥6 mm and CAL ≥5 mm. They had also ≥50% alveolar bone loss in at least two quadrants that was commensurate with amount of plaque accumulation.

All clinical measurements, GCF and plaque sampling and also all treatments were performed by the same calibrated and trained investigator (B.A.) in a standardized manner (single-blinded trial).

Treatment

At the baseline visit, immediately before non-surgical periodontal treatment GCF and subgingival plaque samples were collected per patient and clinical periodontal measurements were recorded in all groups.

In the Q-SRP group, SRP were started with the upper right quadrant, and continued clockwise over four visits at 1-weekly intervals. SRP was performed using an assortment of manual periodontal curettes. The tooth surfaces were instrumented until it was visually and tactilely free of all deposits. Re-examinations were performed at 1, 3 and 6 months following the completion of the SRP in the lower right quadrant.

In the FMUD group, subgingival debridement were performed in upper jaw at the morning session and in lower jaw at the afternoon session. The procedure was completed in two visits (with 45 minutes interval) of the same day. All periodontal sites were debrided by a piezoceramic ultrasonic instrument with unmodified and modified inserts. Each tooth was instrumented till the root surface was visually and tactilely clean and smooth. Re-examinations were performed at 1, 3 and 6 months after the completion of full-mouth subgingival debridman.

In the FMD group, ultrasonic subgingival debridman was combined with an intensive antimicrobial regime with chlorhexidine. The dorsum of the tongue was brushed with a 1% chlorhexidine gel for 1 minute. The patients were instructed to rinse twice for 1 minute with 0.2% chlorhexidine solution. Each tonsil was sprayed four times with 0.2% chlorhexidine spray. Repeated subgingival irrigation with 1% chlorhexidine gel of all the pockets by a blunt needle. This subgingival application was repeated at day 8. For 14 days after the treatment, the patients were instructed to rinse twice daily for 1 min with 0.2% chlorhexidine solution and to spray the tonsils twice daily with a 0.2% chlorhexidine spray. Re-examinations were performed at 1, 3 and 6 months after 14 days washout period.

Standard oral hygiene instructions were given all patients immediately after the first session of Q-SRP group and the completion of the ultrasonic debridement in FMD and FMUD groups.

GCF sampling

GCF was sampled from the buccal aspects of two nonadjacent interproximal sites in single-rooted teeth with PD ≥6 mm, CAL ≥5 mm and visible signs of inflammation at baseline. Re-sampling was repeated at 1 month, 3 months and 6 months after treatment. Standardized filter paper strips were used for GCF sampling. The absorbed fluid volume was measured with a precalibrated electronic device. The paper strips were stored at -40◦C for further analysis.

Subgingival plaque sampling

After GCF collection, the subgingival plaque samples were collected from one site per quadrant with the deepest PD (≥5 mm) of single-rooted teeth at baseline and 1 month, 3 months and 6 months after treatment using two standardized no.30 sterile paper points. Pooled samples were stored at -40◦C for further analysis.

Measurement of calprotectin, osteocalcin, and NTx levels in GCF.

Two paper strips were pooled, placed in 300 microliter phosphatebuffered saline containing 0.05% polysorbate 20 and incubated for 20 minutes at room temperature on an orbital shaker. The fluid from the paper strips was recovered by centrifugation at 13 000 rpm for 5 minutes at +4◦C. Calprotectin, osteocalcin and NTx levels in GCF samples were measured by ELISA using commercial kits in line with the manufacturer's guidelines. Plates were measured at 450 nm with 650 nm as a reference wavelength by using an ELISA reader. Cytokine concentrations were calculated from the standard curve. GCF results were expressed as total amounts at two sites per sampling time.

Molecular detection of target bacteria in subgingival plaque.

The quantitative real-time PCR (qPCR) method was used for the quantitative detection of the selected periodontal pathogens in subgingival plaque. Total microbial DNA on paperpoints were extracted using High Pure PCR Template Preparation Kit according to the manufacturer's instructions. LightCycler FastStart DNA Master SYBR Green I Kit was used for the detection of target oligonucleotides according to the manufacturer's instructions. Real-time PCR protocols were constructed considering specific primer sets. Standard curves were constructed for the calculation of target oligonucleotides in the samples.

Statistical Analysis

Repeated measures analysis of variance (ANOVA) was used to detect intra- and intergroup differences in clinical, microbiological and biochemical parameters. Analysis of covariance was used for comparison of the improvements in parameters between the three treatment groups adjusting for the potential difference in the baseline data and Bonferroni test was performed post-hoc. The significance level was set at p <0.05.